VTA-SNr Dopaminergic Projection Mediates Chronic Social Defeat Stress-induced Locomotor Decline And Parkinson's Disease Vulnerability

Background: Midbrain dopamine neurons play a key role in regulating the behavior,such as voluntary movement,reward processing,and working memory.Two neighboring nuclei,the substantia nigra pars compacta(SNpc))and the ventral tegmental area(VTA)have the largest populations of these neurons.The dysfunction of these dopamine neurons has been associated with the development of mental disorder such as depression and neurodegenerative diseases such as parkinson's disease.However,few studies have focused on the interaction between the dopamine neurons in these two areas.Major depressive disorder is a debilitating disease characterized by diverse symptoms,including low mood,thinking retardation and psychomotor inhibition.The dysfunctions of VTA dopamine neurons were found in both patients and the animal model of depression.While the mice subjected to chronic social defeat stress show the obvious depression-like behaviors,such as anhedonia,anxiety depression and social avoidance,few studies on the changes of motor function in mice.CSDS can induce the change in the firing of VTA-DA neurons.Furthermore,the regulation of the activity and specific neural projection of these neurons impact the depression-like behavior induced by CSDS.However,there is still a need for further study on the relationship between the psychomotor inhibitory behavior and VTA dopamine neurons in the CSDS mice.Depression is the most common complication of parkinson's disease(PD).Few studies have suggested depression as a risk factor for pd susceptibility.PD is a neurodegenerative disease characterized by muscle tremors,muscle rigidity,and decreased mobility,with the degeneration of SNpc-DA neurons as the pathological hallmark.Using the neurotoxin,1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP),selective destruction of SNpc-DA neurons can be triggered,resulting in a PDlike syndrome,which widely used in the pd studies.In this study,mice suggested to csds were injected MPTP by intraperitoneal,which study the effect of stress on the pathological phenomena caused by MPTP? Methods: 1.Mice exposed to CSDS were subjected to the spontaneous locomotor activity test(LMT),the social interaction test(SIT),the forced swimming test(FST),the rotarod test(RT)and the grip strength test(GST).2.CSDS mice of chemogenetic activation of VTA dopamine neurons,or the neurons projectiong to snr were subjected to the spontaneous locomotor activity test.3.The MED64 was used to record the firing changes in VTA dopaminergic neurons activated by chemogenetics.4.Immunofluorescence was used to validate infection effeciency of AAV in the VTADA neurons and recover the types of neurons projected by VTA dopaminergic neurons in SNr 5.Viral anterograde tracer was used to map the VTA-DA output and show the difference of the output in CSDS mice from in Control mice.6.Optogenetics activated the VTA-DA neurons projecting to SNr in phasic or tonic.7.Viral anterograde and retrograde tracer was used to depict the region innervated by VTA-DA neurons and the neurons projecting to SNpc-DA neurons in SNr.8.Viral anterograde,anterograde trans-synaptic and retrograde tracer was used to depict the local circuits between the neurons projected from VTA and to SNpc-DA neurons in SNr.9.CSDS mice were injected MPTP by intraperitoneal.Results: 1.After CSDS,defeated mice showed robust depression-like behaviors in the social interaction test and the forced swimming test.The spontaneous motor ability of CSDS mice had significantly lower than control mice in the spontaneous locomotor activity test.But no significant differences between CSDS and control mice were observed in the rotarod test and grip strength test.2.A virus containing h M3 Dq was bilaterally injected into the VTA.Infection effeciency of virus in the VTA-DA neurons of wildtype mice was validated from brain slices.The activation of VTA-DA neurons firing by chemogenetics was recorded by MED64.The activating group had more spontaneous locomotor activity compared with the nonactivating group.3.Mice were injected viral anterograde tracer to determine the identities of the VTADA neuronal projections altered by CSDS.Representative images of EYFP-positive terminals in the SNr are shown.Moreover,the immunofluorescence staining resulshowed that the representative image indicated the GAD67-positive neurons were surrounded by EYFP-positive terminals within SNr region.The VTA-SNr outputs were significantly decreased in CSDS.Analysis of the labelled projections from VTA-DA neurons in other brain regions,comprised of long-range outputs,revealed significant differences of PAG between the control and CSDS groups,while no differences of rest brain regions.4.Mice were given stereotaxic injections of the retrograde adeno-associated virus with ChR2 into the mSNr.Selectively phasic optogenetic-stimulation of the VTA-SNr circuit causes the increased spontaneous locomotor activity,while Tonic is no change.5.Mice were given stereotaxic injections of the retrograde adeno-associated virus with hM3Dq into the m SNr.Infection effeciency of virus in the VTA-DA neurons projecting to m SNr of wildtype mice was validated from brain slices.The activation of these neurons firing by chemogenetics was recorded by MED64.The activating group had more spontaneous locomotor activity compared with the non-activating group.6.The mice exposed to the 10-day stress and consequently received an intraperitoneal injected of MPTP before perfused.With immunofluorescence technology(TH staining),the CSDS+MPTP mice exhibited a significant reduce in the number of SNpc-DA neurons compared with the Control+NS and the Control+MPTP group,as assessed by counting the number of TH-positive cells in SNpc region.7.Cell-type-specific anterograde tracing and retrograde trans-synaptic tracing revealed VTA-DA terminals that were predominantly located in the medial region of the SNr and an overwhelming number of neurons projecting to SNpc-DA neurons in the lateral part of the SNr.The anterograde transsynaptic tracing were used to further decipher a local circuit in the medial and the lateral of SNr region.Conclusions: 1.Chronic social defeat stress affects the spontaneous locomotor activity of mice greatly,but showed little influence on the passive motor ability.2.Targeted activation of VTA-DA neurons rescued the CSDS-induced impairment of spontaneous locomotor activity.3.CSDS altered the VTA-DA neuronal projections in the Whole-brain.VTA-DA neurons projected to SNr-GABA neurons,and the VTA-SNr outputs were significantly decreased in CSDS.4.Selectively phasic,not Tonic,optogenetic-stimulation of the VTA-SNr circuit causes the increased spontaneous locomotor activity.5.Chemogenetic activation of VTA-SNr projections reversed CSDS-induced impairment of spontaneous locomotor activity.6.The CSDS induced the vulnerability of SNpc neurons in to Parkinson's disease toxin MPTP.7.There is a multiple-synaptic pathway from the VTA-DA through the local circuit of the SNr to SNpc-DA neurons,which mediates the CSDS-induced increasing risk of Parkinson's disease.